PstI
PstI is a type II restriction endonuclease isolated from the Gram negative species, Providencia stuartii.
Function
PstI cleaves
Recognition Sequence | Cut Site |
---|---|
5'CTGCAG 3' 3'GACGTC 5' |
5'--CTGCA G--3' 3'--G ACGTC—5' |
The PstI
PstI is functionally equivalent to BsuBI. Both enzymes recognize the target sequence 5'CTGCAG. The enzyme systems have similar methyltransferases (41% amino acid identity), restriction endonucleases (46% amino acid identity), and genetic makeup (58% nucleotide identity).[6] These observations suggest a shared evolutionary history.
When examining the preferential double strand cleavage of DNA, the restriction endonuclease PstI bind to pSM1 plasmid DNA.[7]
DNA cloning
PstI is a useful enzyme for DNA cloning as it provides a selective system for generating hybrid DNA molecules.[8] These hybrid DNA molecules can be then cleaved at the regenerated PstI sites. Its use is not limited to molecular cloning; it is also used in restriction site mapping, genotyping, Southern blotting, restriction fragment length polymorphism (RFLP) and SNP.[9] It is also an isoschizomer restriction enzyme SalPI from Streptomyces albus P.[10]
Cleavage
PstI preferentially cleaves purified pSM1 DNA without being influenced by the superhelicity of the substrate.[11] However, it is not known whether the effects of this cleavage occurs upon binding to the recognition site or DNA scission. Its differential cleavage rates at different restriction sites is due to the five features of duplex structure. The proximity to the ends in linear DNA molecule, variation in DNA sequence within the recognition sites for enzymes, short distance between regions of unusual DNA sequences and recognition sites, and lastly the special structures such as loops and hairpins. The collective effect of these five factors could affect the accessibility of the restriction enzyme to its recognition sites.
Relation
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